The long term goal of our program is to develop novel broad spectrum chemotherapeutics that can be used for Phase 1 clinical trials for treating the diseases caused by priority pathogens such as Francisella tularensis, Burkholderia pseudomallei and Yersinia pestis. We have developed a series of high affinity inhibitors of the fatty acid biosynthesis enoyl reductase (Fabl) enzyme that demonstrate in vitro potency against F. tularensis, M. tuberculosis and B. pseudomallei, and are active in an animal model of tularemia, confirming the validity of the target and of our approach. The goal of the present project is to dramatically improve the therapeutic dose by determining how factors such as formulation and modes of delivery, and structural modifications impact the in vivo antibacterial activity of the compounds. Studies will initially focus on the category A pathogen F. tularensis and the tularemia model of infection as this is the system in which we have had the most success. As the project evolves the information gained will be used to drive additional studies on B. pseudomallei and the melioidosis animal model of infection and Y. pestis. We will perform lead optimization studies on the Fabl inhibitors to: (i) improve pharmacology and reduce metabolic liability, (ii) facilitate deliverability/bioavailability, (iii) improve release kinetics, and (iv) assess alternative routes of delivery to optimize candidates for preclinical studies and performing required benchmarks for submission of an IND application. Our goal of developing inhibitors with enhanced in vivo efficacy will be achieved with the following Specific Aims: Aim 1: Effect of formulation and method of delivery on PK and in vivo antibacterial activity. Aim 2: Introduction of structural modifications to improve PK/PD. Aim 3: Extension to Other Priority Pathogens. RELEVANCE (See instructions): This proposal is to enhance the therapeutic effect of broad spectrum chemotherapeutics with efficacy against F. tularensis and B. pseudomallei and Y. pestis infections. In addition, such chemotherapeutics can be used to treat bacterial agents with significant health relevance, particularly Gram-positive pathogens, including MRSA and Gram-negative pathogens including B. cenocepacia, A. baumannii, and P. aeruginosa